Figure 1: Human ENTH forms a stable hexamer with a predicted membrane-binding interface, the bound PIP2 molecules are all located on one side of the protein complex.
The Garcia Alai team develops methods for sample optimization and characterization for SAXS, MX and EM experiments and applies systematic pipelines of biophysical techniques to solve dynamic structural puzzles with particular focus on protein-lipid interactions.
Previous and current research
Epsin and Sla2 form assemblies through phospholipid interfaces
In clathrin-mediated endocytosis, adapter proteins assemble together with clathrin through interactions with specific lipids on the plasma membrane. We have shown that the membrane-proximal domains ENTH of epsin and ANTH of Sla2 form complexes by sharing phosphatidylinositol 4,5-bisphosphate (PIP2) lipid molecules. The crystal structures of epsin Ent2 ENTH domain from S. cerevisiae in complex with PIP2 and Sla2 ANTH domain from C. thermophilum illustrate how allosteric phospholipid binding occurs. A comparison with human ENTH and ANTH domains reveal that only the human ENTH domain can form a stable hexameric core in presence of PIP2, which could explain functional differences between fungal and human epsins.
Future projects and goals
In collaboration with groups working in the membrane protein field we are developing highly automated pipelines for sample preparation. These will integrate membrane solubilisation, detergent screening, lipid reconstitution and sample optimization in an automated workflow. Funded access to these new pipelines will be available through the EC-funded iNEXT project.